• Archives of Pharmacal Research
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• v.16 no.1
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• pp.57-61
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• 1993

Based on the turbidimetric response of protein with 50% trichloroacetic acid (TCA), this study aims to introduce an assay method for protein in solution. The standard procedure consists of mixing equal volume of sample solution (standard or unknown) with 50%-TCA solution and measuring the absorbance at 450 nm after 20 min. The absorbances of the solutions were almost stable over 120 min at room temperature. This assy method is simple, reproducible, and tolerant to many interfering substances. It can detect less amount than $10\mu$g/ml of bovin serum albumin. The assay method has low protein-to-protein variability over wide range of molecular weight.

• Biotechnology and Bioprocess Engineering:BBE
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• v.9 no.4
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• pp.241-244
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• 2004

Protein A molecular thin film was fabricated as a platform of antibody-based biosensor. For the immobilization of the protein A thin film, a viologen multilayer was built up using the Langmuir-Blodgett (LB) technique, and then, protein A was adsorbed on the viologen LB film by an electrostatic interaction force, which was formed as a hetero-film structure. For the deposition of viologen, surface pressure area ($\pi$-A) isotherm was investigated. The fabricated protein A-viologen hetero LB film was investigated using atomic force microscopy (AFM). Using the developed molecular film, antibody immobilization and fluorescence measurement was carried out.

• Korean Journal of Plant Tissue Culture
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• v.27 no.4
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• pp.283-297
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• 2000

Seeds are an ideal repository for recombinant proteins in molecular farming applications. However, in order to use plant seeds efficiently for the production of such proteins, it is necessary to understand a number of fundamental biological properties of seeds. This includes a full understanding of promoters which function in a seed-specific manner, the subcellular targeting of the desired polypeptide and the final form in which a protein is stored. Once a biologically active protein has been deposited in a seed, it is also critical that the protein can be extracted and purified efficiently. In this review, these issues are examined critically to provide a number of approaches which may be adopted for production of recombinant proteins in plants. Particular attention is paid to the relationship between subcellular localization and protein extraction and purification. The robustness and flexibility of seed-based production is illustrated by examples close to or already in commercial production.

• Journal of the Korean Magnetic Resonance Society
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• v.20 no.1
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• pp.22-26
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• 2016

FANCJ is a DNA helicase which contributes genome stability by resolving G-quadruplex DNA from 5' to 3' direction. In addition to main ATPase helicase core, FANCJ has the protein binding region at its C-terminal part. BRCA1 and BLM are the binding partner of FANCJ and these protein-protein interactions contribute genomic stability and the proper response to replication stress. As the first attempt for studying FANCJ-BLM interaction, we prepared BLM binding region of FANCJ and characterized with CD and NMR spectroscopy. FANCJ (881-941) with N-ter 6xHis was purified as the oligomer. Secondary structure prediction based on CD data revealed that FANCJ (881-941) composed with ${\beta}$ sheet, turn and coils.$^1H-^{15}N$ HSQC spectra showed nonhomogeneous peak intensities with less number of peaks comparing than the number of amino acids in the construct. It indicated that optimization should be necessary for detailed further structural studies.

• Bulletin of the Korean Chemical Society
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• v.26 no.3
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• pp.413-417
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• 2005

Fluorescence of green fluorescent protein mutant, 2-5 GFP is observed during denaturation by guanidine. The fluorescence intensity decreases exponentially but the fluorescence lifetime does not change during denaturation. The fluorescence lifetime of the denatured protein is shorter than that of native form. As the protein structure is modified by guanidine, solvent water molecules penetrate into the protein barrel and protonate the chromophore to quench fluorescence. Most fluorescence quenchers do not affect the fluorescence of native form but accelerate the fluorescence intensity decay during denaturation. Based on the observations, a simple model is suggested for the structural change of the protein molecule during denaturation.

• Proceedings of the Korean Society for Bioinformatics Conference
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• 2000.11a
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• pp.28-31
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• 2000

We describe a hidden Markov model, HMMTIR, for general protein sequence based on the I-sites library of sequence-structure motifs. Unlike the linear HMMs used to model individual protein families, HMMSTR has a highly branched topology and captures recurrent local features of protein sequences and structures that transcend protein family boundaries. The model extends the I-sites library by describing the adjacencies of different sequence-structure motifs as observed in the database, and achieves a great reduction in parameters by representing overlapping motifs in a much more compact form. The HMM attributes a considerably higher probability to coding sequence than does an equivalent dipeptide model, predicts secondary structure with an accuracy of 74.6% and backbone torsion angles better than any previously reported method, and predicts the structural context of beta strands and turns with an accuracy that should be useful for tertiary structure prediction. HMMSTR has been incorporated into a public, fully-automated protein structure prediction server.

• Journal of Pharmaceutical Investigation
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• v.34 no.5
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• pp.369-377
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• 2004

The objective of this study was to investigate the patterns of protein leaching to an external phase during an ethyl acetate-based, double emulsion microencapsulation process. An aqueous protein solution (lactoglobulin, lysozyme, or ribonuclease; $W_1$) was emulsified in ethyl acetate containing poly-d,l-lactide-co-glycolide 75:25. The $W_1/O$ emulsion was transferred to a 0.5% polyvinyl alcohol solution saturated with ethyl acetate $(W_2)$. After the double emulsion was stirred for 5, 15, 30, or 45 min, additional 0.5% polyvinyl alcohol $(W_3)$ was quickly added into the emulsion. This so-called quenching step helped convert emulsion microdroplets into microspheres. After 2-hr stirring, microspheres were collected and dried. The degree of protein leaching to $W_2$ and/or $W_3$ phase was monitored during the microencapsulation process. In a separate, comparative experiment, the profile of protein leaching to an external phase was investigated during the conventional methylene chloride-based microencapsulation process. When ethyl acetate was used as a dispersed solvent, proteins continued diffusing to the $W_2$ phase, as stirring went on. Therefore, the timing of ethyl acetate quenching played an important role in determining the degree of protein microencapsulation efficiency. For example, when quenching was peformed after 5-min stirring of the primary $W_1/O$ emulsion, the encapsulation efficiencies of lactoglobulin and ribonuclease were $55.1{\pm}4.2\;and\;45.3{\pm}7.6%$, respectively. In contrast, when quenching was carried out in 45 min, their respective encapsulation efficiencies were $39.6{\pm}3.2\;and\;29.9{\pm}11.2%$. By sharp contrast, different results were attained with the methylene-chloride based process: up to 2 hr-stirring of the primary and double emulsions, less than 5% of a protein appeared in $W_2$. Afterwards, it started to partition from $W_1\;to\;W_2/W_3$, and such a tendency was affected by the amount of PLGA75:25 used to make microspheres. Different solvent properties (e.g., water miscibility) and their effect on microsphere hardening were to be held answerable for such marked differences observed with the two microencapsulation processes.

• Journal of Applied Biological Chemistry
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• v.48 no.3
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• pp.120-126
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• 2005

Endothelial nitric oxide synthase (eNOS) plays a critical role in vascular biology and pathophysiology. Its activity is regulated by multiple mechanisms such as calcium/calmodulin, protein-protein interactions, sub-cellular locations and phosphorylation at various sites. Phosphorylation of eNOS-Ser1177 (based on mouse sequence) has been identified as an important mechanism of eNOS activation. However, signaling pathway leading to it phosphorylation remains controversial. The regulation of eNOS-Ser1177 phosphorylation by Src and protein kinase A (PKA) was investigated in the present study using cultured mouse aorta endothelial cells. Expression of a constitutively active Src mutant in the cells enhanced phosphorylation of eNOS and protein kinase B (Akt). The Src-stimulated phosphorylation was not attenuated by the expression of a dominant negative PKA regulatory subunit. Neither activation nor inhibition of PKA activity had any significant effect on tyrosine phosphorylation of activation or inactivation site in Src. Based on the results of this study, it is suggested that Src/Akt pathway and PKA signaling may regulate eNOS phosphorylation independently. The existence of multiple mechanisms for eNOS phosphorylation may guarantee endothelial nitric oxide production in various cellular contexts which is essential for maintenance of vascular health.

• Nutritional Sciences
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• v.9 no.4
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• pp.240-247
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• 2006

In this study we investigated the neuroprotective, antioxidative, and hypocholestrolemic effects of dietary soy protein and soy isoflavone in a rat focal brain ischemia model. Weaning Sprague-Dawley rats were fed a 20% casein-based diet (CA), 20% soy protein-based diet (SP), or 0.2% soy isoflavones-supplemented diet (ISO) for 6 weeks. The cortical infarction volume of the ISO group was significantly lower than that of the SP group. The thiobarbituric acid reactive substances (TBARS) were considerably lower in the ISO group than the CA group. Glutatbione peroxidase activities of the SP group were notably higher than those of the CA group. Acetylcholinesterase (AchE) activities of the SP group were significantly decreased compared to the CA group. LDL cholesterol levels and LDL/HDL ratios of the ISO group were lower than those of the CA and SP groups. Our results collectively suggest that soy isoflavones may contribute to neuroprotection by reducing the TBARS and serum LDL/HDL ratio, whereas soy protein may be associated with the regulation of cognitive functions by modulating AchE activity.

• The Journal of the Korea Contents Association
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• v.8 no.1
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• pp.259-271
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• 2008

We propose a protein function finding algorithm that is able to predict specific molecular function for unannotated proteins through domain analysis from protein-protein network. To do this, we first construct protein-protein interaction(PPI) network in Saccharomyces cerevisiae from MIPS databases. The PPI network(proteins; 3,637, interactions; 10,391) shows the characteristics of a scale-free network and a hierarchical network that proteins with a number of interactions occur in small and the inherent modularity of protein clusters. Protein-protein interaction databases obtained from a Y2H(Yeast Two Hybrid) screen or a composite data set include random false positives. To filter the database, we reconstruct the PPI networks based on the cellular localization. And then we analyze Hub proteins and the network structure in the reconstructed network and define structural modules from the network. We analyze protein domains from the structural modules and derive functional modules from them. From the derived functional modules with high certainty, we find tentative functions for unannotated proteins.